Co-culture of osteochondral explants and synovial membrane as in vitro model for osteoarthritis

PLoS One. 2019 Apr 2;14(4):e0214709. doi: 10.1371/journal.pone.0214709. eCollection 2019.

Abstract

The purpose of the current study was to establish an in vitro model for osteoarthritis (OA) by co-culture of osteochondral and synovial membrane explants. Osteochondral explants were cultured alone (control-1) or in co-culture with synovial membrane explants (control-2) in standard culture medium or with interleukin-1β (IL1β) and tumor necrosis factor (TNFα) added to the culture medium (OA-model-1 = osteochondral explant; OA-model-2 = osteochondroal-synovial explant). In addition, in OA-model groups a 2-mm partial-thickness defect was created in the centre of the cartilage explant. Changes in the expression of extracellular matrix (ECM) genes (collagen type-1 (Col1), Col2, Col10 and aggrecan) as well as presence and quantity of inflammatory marker genes (IL6, matrix metalloproteinase-1 (MMP1), MMP3, MMP13, a disintegrin and metalloproteinase with-thrombospondin-motif-5 (ADAMTS5) were analysed by immunohistochemistry, qPCR and ELISA. To monitor the activity of classically-activated pro-inflammatory (M1) versus alternatively-activated anti-inflammatory/repair (M2) synovial macrophages, the nitric oxide/urea ratio in the supernatant of osteochondral-synovial explant co-cultures was determined. In both OA-model groups immunohistochemistry and qPCR showed a significantly increased expression of MMPs and IL6 compared to their respective control group. ELISA results confirmed a statistically significant increase in MMP1and MMP3 production over the culturing period. In the osteochondral-synovial explant co-culture OA-model the nitric oxide/urea ratio was increased compared to the control group, indicating a shift toward M1 synovial macrophages. In summary, chemical damage (TNFα, IL1β) in combination with a partial-thickness cartilage defect elicits an inflammatory response similar to naturally occurring OA in osteochondral explants with and without osteochondral-synovial explant co-cultures and OA-model-2 showing a closer approximation of OA due to the additional shift of synovial macrophages toward the pro-inflammatory M1 phenotype.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chondrocytes / cytology*
  • Chondrocytes / metabolism
  • Chondrocytes / pathology
  • Coculture Techniques
  • Collagen Type I / genetics
  • Collagen Type I / metabolism
  • Horses
  • Interleukin-1beta / pharmacology
  • Interleukin-6 / genetics
  • Matrix Metalloproteinase 1 / genetics
  • Matrix Metalloproteinase 1 / metabolism
  • Models, Biological*
  • Nitric Oxide / metabolism
  • Osteoarthritis / pathology*
  • Synovial Membrane / cytology*
  • Synovial Membrane / metabolism
  • Tumor Necrosis Factor-alpha / pharmacology
  • Urea / metabolism

Substances

  • Collagen Type I
  • Interleukin-1beta
  • Interleukin-6
  • Tumor Necrosis Factor-alpha
  • Nitric Oxide
  • Urea
  • Matrix Metalloproteinase 1

Grants and funding

The current research was funded by the Bright Sparks program of the University of Veterinary Medicine Vienna, Austria to EH. The funding source was not involved in any part of the study, neither the study design, execution of the in vitro experiments, sample analysis, data analysis nor writing of the manuscript or publication.